Sourcing bacterial strains for rum fermentations

[Classic Lloyd]

13 hours ago, Silk City Distillers said:

Probably one of the best recent PhD dissertations I've read recently on this exact topic:

 The microbial ecology of a rum production process

Dr. Victoria Green - Working at Bundaberg Rum in Australia.

Thanks for sharing this - it's fantastic. We're currently running a series of fermentations with a culture cultivated from fresh-pressed sugarcane, which have over a few generations, has whittled down to one dominant yeast, with 1-2 other yeast species also in there.  I'd assumed that the dominant species was S. cerevesiae, but this article makes me wonder if I'm not working with something different.

[Simpo]

On 1/24/2019 at 9:07 AM, Classic Lloyd said:

...I'd assumed that the dominant species was S. cerevesiae, but this article makes me wonder if I'm not working with something different.

Have you found out since? 

[Curators Reserve]

60,000,000 litres / year output of 96-99% ABV Neutral Cane Spirit from the largest Molasses based Distillery in Australia - https://wilmarbioethanol.com/

2 x 1,000,000L (ML) Continuously run FV system - alternating back & forth and is the BioStil Process that is utilised in Brazil & India.

but what there's more..

100% S. Pombe culture - a ~30%Brix vinasse output results from the focus  to minimise the actual water intake/ demand for the operation of the system and the Pombe provides more 'optimal' stability over 5-6 day ferment in those volumes versus S.cerevesiae.

Yeast cake is consistently removed to manage and control yeast cell health & viability.

For any Aussie out there reading this - the Yeast Cake from this distillery was also the Vegemite yeast extract production site between 1951 to 1971 - aside from CUB in Melbourne.

 

[Silk City Distillers]

Wasn't Vegemite historically just spent brewers yeast extracts?

[Curators Reserve]

Yes - CUB is Carlton United Breweries - historically the largest Brewery group in Australia.

This Molasses Ethanol Distillery was next biggest yeast production site & apparently produced a superior yeast extract for the day - not S. Pombe.

Anyone used Aspergillus in a molasses/ rum wash to push Citric and Malic acid output rather than Lacto?

Just reading recently also that the sugarcane bagasse & more specifically the cane wax/ external cuticle layer is high in C16 & C18 fatty acids..

Within the end-to-end process sugarcane mill & distillery facility that I work in - I have noted the 'rum oils' many times as a distinct surface sheen/ slick when clarified juice is cooled allowing the oils to coalesce.

[Silk City Distillers]

There are plenty of other plant based sources of long-chain fatty acids and waxes that are fairly similar, depending on what your target is.

Depending on what you want to target, it's fairly easy to find a plant-based match.  You can even engineer a specific profile using GRAS ingredients.

For example, if I want to target C12-C16, I might use a refined coconut oil (no, you don't get any coconut flavor or aroma in the distillate).

If I want a high percentage of C16 - Rice Bran Oil.

If I want to target C18 - Almond Oil.

C16-C18 - Peanut Oil.

If I want a higher percentage of waxes - Beeswax, or heck, just go with cane waxes as well.

Folks who mash corn high in fatty acids see similar oil-slicks - corn oil high in C10 and C16.

Probably more interesting are the long-chain wax esters, meaning you'd probably need two components to match.  Like Peanut Oil and Beeswax.

Which one of these makes the delicious rum oil?  Which one of these tastes terrible?

[bostonapothecary]

Y'all may enjoy this paper on "bauer oil" that turned up while translating Kervegant's 1946, Rhum and Canne Eau-de-vie.

Bauer oil may be a match for Arroyo's "rum oil" and these ideas go all the way back to Micko and then across to old authors that talked of Cognac oil. The "oil" is a complex mixture and has both saponifiable and non-saponifiable parts. The bulk of it is saponifiable. The portion that isn't may be the realm of rose ketones and all that nonsense.

In my experience with the birectifier, when concentrated, these fractions aren't harmonious and can seem intensely acrid, but when diluted to normal levels and part of a sensory matrix that includes ethyl acetate they define a spirit and represent quality.

One thing recently learned about Pombe fission yeasts is that they have on average much thicker cell walls than budding yeasts. This gives their lees a lot more potential for aroma when broken down. In a typical ferment, fission yeasts also have the tendency to produce more esters because the cell wall at the point of their division is extra thick and a portion dissolves into the ferment upon successful separation.

[Classic Lloyd]

15 hours ago, Simpo said:

Have you found out since? 

That strain was definitely a S. Cerevesiae. It eventually adapted to be a prolific producer of acetaldehyde and we ended up shelving it.  Generally speaking, our experience is that wild strains lose a lot of their dynamic nature (and diversity) in just a few generations.

[Classic Lloyd]

3 hours ago, Curators Reserve said:

Anyone used Aspergillus in a molasses/ rum wash to push Citric and Malic acid output rather than Lacto?

Is this a thing? I work occasionally with aspergillus and have found it to be very finicky on anything but grains. (I suppose you could add inoculated grains, but then it would cease being a rum)

[Curators Reserve]

Commercial Citric acid production is typically Apergillus in a molasses wash ferment.

Malic Acid is also produced by Aspergillus..

Aside from inherent LAB in freshly crushed juice - ester formation from increased Citric & Malic acid output from Aspergillus is of interest..

 

[JustAndy]

On 9/30/2019 at 6:26 AM, Classic Lloyd said:

Is this a thing? I work occasionally with aspergillus and have found it to be very finicky on anything but grains. (I suppose you could add inoculated grains, but then it would cease being a rum)

There is a style of shochu made with koji-rice and sugar (kokuto shocho), as well a sort of similar process for batavia Arrack made with molasses and an inoculated rice starter. 

[Silk City Distillers]

On 9/30/2019 at 9:15 AM, bostonapothecary said:

In my experience with the birectifier, when concentrated, these fractions aren't harmonious and can seem intensely acrid, but when diluted to normal levels and part of a sensory matrix that includes ethyl acetate they define a spirit and represent quality.

 

Not uncommon when talking about flavor and aroma compounds, where an increase in concentration can take a taste or smell from pleasant to awful, or worse.  That creates a whole other level of complexity where it simply isn't about creation of a specific compound as part of the process, but the creation of that compound at a specific concentration.

[Classic Lloyd]

On 10/1/2019 at 11:56 AM, JustAndy said:

There is a style of shochu made with koji-rice and sugar (kokuto shocho), as well a sort of similar process for batavia Arrack made with molasses and an inoculated rice starter. 

Minor update - this idea has stuck with me and I'm running a test fermentation to see if I can get Aspergillus Oryzae to grow directly on sugar cane (as opposed to a grain), and it seems to be taking so far. One curveball - the sugar cane I'm using had a red/purple colored fungus that was present on the husk (and slightly in the substrate), so we'll see if that sticks around.

[Curators Reserve]

Great stuff - and your Aspergillus culture source?

There is good stuff on google Scholar about ligno-cellulosic R&D in this..

http://docs.bvsalud.org/biblioref/2018/09/914332/cultivation-of-aspergillus-niger-on-sugarcane-bagasse-with-vinasse.pdf

What do you typically do with your bagasse otherwise or have you sourced some cane sticks to trial?

[Silk City Distillers]

Red sugarcane fungus - https://www.longdom.org/open-access/a-review-on-red-rot-the-cancer-of-sugarcane-2157-7471-1000S1-003.pdf

 

[Silk City Distillers]

I still don't really understand the point of Aspergillus for rum production.  It's a necessary component of lots of asian spirit types due to the fact that their starch sources generally aren't malted - sweet potato, rice, etc.  Without Aspergillus, there would be no sugar for yeast to ferment - absolutely ingenious - absolutely necessary (as in, no other choice).

What's the goal for rum?  Many of the flavor compounds produced by Aspergillus aren't volatile (acids for example), not to mention the high potential for funky off-flavor and aroma of koji ferments.  Shochus a good reference point for typical koji profiles, bad shochu a really good example of atypical off flavors that would ruin rum (bitter pickles).  Keep in mind cultural expectations of spirit types.  Sure, there are some interesting compounds in there, get that, but there are also flavor profiles that are pretty foreign to western palates.

Also keep in mind koji production, generally using sterile steamed rice.  With cane, you aren't really going to be sterilizing it with heat for koji production.  You are going to get a host of bacterial and fungal components growing, likely good and bad.  You could grow some really nasty fungus here.  I suppose you could chop some cane, roast it to sterilize, use it as a koji starter base without the massive microbial impacts, but go back to my original point, why?

On 10/1/2019 at 12:56 PM, JustAndy said:

There is a style of shochu made with koji-rice and sugar (kokuto shocho),

Imagine the brown sugar base would make for a very rum-like profile, probably far milder in flavor than typical Shochu - less koji, less rice.  Interesting history on that, but has more to do with access to sugar as a result of sugar production, and taking advantage of it.  This is more like an interesting cultural mashup than some sort of symbiotic fermentation product though.  You could argue, kind of similar to home distillers goosing their product yield by adding sugar to simple grain fermentations.

Played around with this a bit, as we have a big local Korean population - who always ask about soju.  I felt you could get a far milder (aka western acceptable) flavor profile using all enzyme vs. koji.  Yeah I know, purists rolling in their graves.

[Classic Lloyd]

On 10/20/2019 at 2:20 AM, Curators Reserve said:

Great stuff - and your Aspergillus culture source?

There is good stuff on google Scholar about ligno-cellulosic R&D in this..

http://docs.bvsalud.org/biblioref/2018/09/914332/cultivation-of-aspergillus-niger-on-sugarcane-bagasse-with-vinasse.pdf

What do you typically do with your bagasse otherwise or have you sourced some cane sticks to trial?

Ahhhh - thanks for sharing this article! At first glance, this should help answer my main question (how will the koji interact with the cane plant structure / fiber?).

This is a test fermentation, so my sugar cane is from the Mexican grocery down the road from me. Â The spores are sourced from a company in Japan (Hishiroku) through a reseller here in the US - I namely use this strain for other non-alcohol uses (shio koji / miso). Â I should also note that I'm using oryzae, not niger for this test, but if the oryzae shows promise, A. niger would be an interesting next step.

[JustAndy]

On 10/20/2019 at 5:08 AM, Silk City Distillers said:

Imagine the brown sugar base would make for a very rum-like profile, probably far milder in flavor than typical Shochu - less koji, less rice.  Interesting history on that, but has more to do with access to sugar as a result of sugar production, and taking advantage of it.  This is more like an interesting cultural mashup than some sort of symbiotic fermentation product though.  You could argue, kind of similar to home distillers goosing their product yield by adding sugar to simple grain fermentations.

 

That's probably true as well, but I assumed that use of koji rice in brown sugar shochu was to provide some yeast nutrients and acid modulation, as sugar can be deficient in both. 

[Silk City Distillers]

Great point, agree.

[Classic Lloyd]

On 10/21/2019 at 1:47 PM, JustAndy said:

That's probably true as well, but I assumed that use of koji rice in brown sugar shochu was to provide some yeast nutrients and acid modulation, as sugar can be deficient in both. 

This is probably part of it, but a lot of the examples I know of use koji (or variant) for their enzymatic power. My assumption is that there would be some sort of benefit to the overall fermentation, either attenuation or additional ester production.  My experiment is around growth on sugar cane; were I to take the next step, I might try and utilize the enzymes, but not the koji, by heating it in a way similar to a whiskey mash killing the koji in the process.  Most of the non-alcoholic uses (e.g. shoyu, miso, fish sauce) of koji do something similar to this through the use of salt in the fermentation 

That said, it looks like the shochu you mentioned and arracks both utilize a sake-style by adding in koji and fermentables periodically throughout the fermentation. I went out and got a bottle of Batavia arrack, and I have to say that I'm really enamored with the result. Short of some of the Clairins I've tried, it might be my favorite agricole-like rum. But....

On 10/20/2019 at 7:08 AM, Silk City Distillers said:

Imagine the brown sugar base would make for a very rum-like profile, probably far milder in flavor than typical Shochu - less koji, less rice.  Interesting history on that, but has more to do with access to sugar as a result of sugar production, and taking advantage of it.  This is more like an interesting cultural mashup than some sort of symbiotic fermentation product though.  You could argue, kind of similar to home distillers goosing their product yield by adding sugar to simple grain fermentations.

I think you're right here. Part of the test on the sugar cane was to see if you could design something more symbiotic, as I suspect the koji is mostly working on the grain separately to create that agricole-like funkiness. Using grains is a fallback for sure, but it'd be great to design something cane / sugar based.

[Silk City Distillers]

I take back my earlier statement, after doing a little research, it's actually far more likely that the rice and koji represent a methodology to store and propagate yeast and bacterial cultures in a sustainable way.  Think along the lines of sourdough starter cultures.

Koji being necessary to hydrolyze starches into sugars to feed the bacteria and yeast.  By sterilizing (steaming) rice prior to inoculating w/ mixed-culture koji - it's equivalent to step starting a liquid yeast culture.  The aspergillus isn't the magic, it's the yeast and other bacteria that exist along side it.  Koji is fairly portable, easily stored.  While not sterile, as a thousand+ year old method of propagating yeast starters, it's a pretty good approach.  

Looking at a few of the current "mainstream" brands - they actually use minimal amounts of rice in their product.  "By the Dutch" Battavia Arak specifically indicates it's used as a culture starter.

Quote

Raw materials:
100% Sugarcane molasses. Small amount of local red rice has been used in the process as trigger/booster to start fermentation (no ingredient).

The challenge is, you can't replicate this using only koji.  The koji is only the enabler here.  The important "stuff" is the other yeast and bacteria co-existing in their starter cultures.  Suppose you'd need to get on a plane and ask to have some.  Think of this as the equivalent of the muck/dunder pit.  You can put fresh dunder in a tank, call it a pit, maybe you'll get some bacterial growth depending on the environment - but without the right bugs, you don't get the right result.

[Silk City Distillers]

Not to mention, there is literature pointing to "red rice koji" in arak potentially being Monascus purpureus, and not Aspergillus sp..

 

[bostonapothecary]

Something to keep in mind is that Batavia Arracks were revealed to use Pombe yeasts at about the turn of the century (by Eijkman who became a Nobel laureate). I have one of their yeasts collected in the 1920's. I have not used this yeast yet, but my research partner (who is way ahead of them) thinks it is the champion among our 14 historic Pombe yeasts.

I collected and translated most all the Pombe literature. I had a few citations that Kervegant did not and he had one major one I haven't found yet. That paper supposedly details how a feature of their fermentations was being very high pH. I have not seen that claim repeated.

I surveyed one Batavia Arrack with the birectifier, but never published the results because it turned up some things I could not explain. One was that the first fractions were abnormally weak relative to any other heavy spirit meaning they were quite low on ethyl acetate. At the time, I thought this may have been due to aggressive cutting to sell the product as a concentrate for the flavor industry. It could have been cut in a way to not pay taxes on the fractions that did not matter to the flavoring industry. That is probably not the case and instead it is sold as a new make spirit and sees fairly low ester creation. At the back end of the spirit, the fraction 5 is abnormally strong on aroma I'd attribute to rum oil/rose ketones, but it is not estery and the very last fractions are not very acidic, but do seem to possess a butyric/butter sour kind of character that is really pleasant. It was also lower than average on fusel oil.

The quality of Batavia Arrack may just be about maximizing the ability of their yeast to create rum oil in a high pH environment similar to what was described by Arroyo.

[bostonapothecary]

8 hours ago, Classic Lloyd said:

I think you're right here. Part of the test on the sugar cane was to see if you could design something more symbiotic, as I suspect the koji is mostly working on the grain separately to create that agricole-like funkiness. Using grains is a fallback for sure, but it'd be great to design something cane / sugar based.

Keep in mind, some of the agricole funkiness can be the "vesouté" aroma described by Kervegant that comes from cane that is not defecated. He called this aroma a flaw, but I enjoy it, particularly in cocktails.

[Silk City Distillers]

Historically, it looks like there was quite a bit of variation in the fermentation stocks used for these "Araks" - Sugar Palm, Coconut Palm Sugar, Cane Juice, Molasses, etc.